(a) Field of the Invention
The present invention relates to a gene being capable of modifying resistance against a heavy metal or salt, or accumulation properties, and a transformant produced using the gene. More particularly, the present invention relates to a gene being capable of modifying resistance against a heavy metal or salt, or accumulation properties, a recombination vector including the gene that improves resistance against salt or drought, a transformant produced using the recombination vector, a transgenic plant that has good resistance against a heavy metal, salt, or drought, efficiently removes or accumulates a heavy metal, and decreases uptake of a heavy metal, a phytoremediation method using the transgenic plants, and a method of developing a safety plant.
(b) Description of the Related Art
Heavy metals are major environmental toxicants, which cause reactive oxidation species generation, DNA damage, and enzyme inactivation by binding to active sites of enzymes in cells.
Contamination of the environment with heavy metals has increased drastically due to industrialization. By the early 1990s, the worldwide annual release had reached 22,000 tons of cadmium, 954,000 tons of copper, 796,000 tons of lead, and 1,372,000 tons of zinc (Alloway B J & Ayres D C 1993 Principles of environmental pollution. Chapman and Hall, London). The soils contaminated with heavy metal inhibit normal plant growth and cause contamination of foodstuffs. Many heavy metals are very toxic to human health and carcinogenic at low concentrations.
Therefore removal of heavy metals from the environment is an urgent issue. Traditional methods of dealing with soil contaminants include physical and chemical approaches, such as the removal and burial of the contaminated soil, isolation of the contaminated area, fixation (chemical processing of the soil to immobilize the metals), and leaching using an acid or alkali solution (Salt D E, Blaylock M, Kumar N P B A, Viatcheslav D, Ensley B D, et al. 1995. Phytoremediation: a novel strategy for the removal of toxic metals from the environment using plants. Bio-Technology 13, 468-74; Raskin I, Smith R D, Salt D E. 1997 Phytoremediation of metals: using plants to remove pollutants from the environment. Curr. Opin. Biotechnol. 8, 221-6). These methods, however, are costly and energy-intensive processes.
Phytoremediation has recently been proposed as a low-cost, environment-friendly way to remove heavy metals from contaminated soils, and is a relatively new technology for cleanup of contaminated soil that uses general plants, specially bred plants, or transgenic plants to accumulate, remove, or detoxify environmental contaminants. The phytoremediation technology is divided into phytoextraction, rhizofiltration, and phytostabilization. Phytoextraction is a method using metal-accumulating plants to extract metals from soil into the harvestable parts of the plants; rhizofiltration is a method using plant roots to remove contaminants from polluted aqueous streams; and phytostabilization is the stabilization of contaminants such as toxic metals in soils to prevent their entry into ground water, also with plants (Salt et al., Biotechnology 13(5): 468-474, 1995).
Examples of phytoremediation are methods using the plants of Larrea tridentate species that are particularly directed at the decontamination of soils containing copper, nickel, and cadmium (U.S. Pat. No. 5,927,005), and a method using Brassicaceae family (Baker et al., New Phytol. 127:61-68, 1994).
In addition, phytoremediation using transgenic plants that are generated by introducing genes having resistant activity for heavy metals have been attempted. Examples of heavy metal resistant genes are AtATM3 (ABC transporters of mitochondria), CAX2 (Calcium exchanger 2), cytochromium P450 2E1, NtCBP4 (Nicotiana tabacum calmodulin-binding protein), GSHII (glutathione synthetase), AtPcr1 (plant cadmium resistance), AtPDR12 (pleiotropic drug resistance), or MRT polya peptide (metal-regulated transporter polypeptide). AtATM3 is an ABC-type transporter that improve cadmium and lead resistance of over-expressed transgenic plants, and increases cadmium contents in transgenic plants (Kim et al., Plant Physiol. 140:922-932, 2006), CAX2 accumulates heavy metals including cadmium and manganese in plants (Hirschi K D et al., Plant Physiol. 124:125-134, 2000), cytochromium P450 2E1 uptakes and decomposes organic materials such as trichloroethylene (TCE) (Doty S L et al., Proc. Natl. Acad. Sci. USA 97:6287-6291, 2000). Transgenic plant transformed with NtCBP4 has resistant activity for nickel (Arazi T et al. Plant J. 20:171-182, 1999), GSHII accumulates cadmium (Liang Zhu Y et al., Plant Physiol. 119:73-80, 1999), plants that over-express AtPcr1 has cadmium resistance by decreasing cadmium content (Song et al., Plant Physiol. 135:1027-1039, 2004, Korean Patent Application No. 2003-0058299 and U.S. patent application Ser. No. 10/907,694), transgenic plants transformed with AtPDR12 improves lead resistance by lead contents in a plant (Lee et al., Plant Physiol. 138:827-836, 2005), and MRT polypeptides remove heavy metals such as iron, cadmium, manganese, and zinc from soils (U.S. Pat. No. 5,846,821.
Recently, it is reported that bacteria and yeast genes as well as plant genes can be introduced into a plant to effectively improve heavy metal resistance and accumulation. For example, merB (organomercurial lyase) was reported to be able to decompose organic mercury materials (Bizily S P et al., Proc. Natl. Acad. Sci. USA 96:6808-6813, 1999). In addition, a ZntA gene, which is a P-type pump of bacteria, was reported to increase resistance against and also, less uptake them when it was over-expressed by merB (Lee et al., Plant Physiol. 133:589-596, 2003; Korean Patent No. 0515520). An Ycf1 (yeast cadmium factor1) gene, which is an ABC-type carrier of yeast, was reported to be expressed in a plant and thereby, increase resistance against cadmium and lead and accumulation thereof when it was over-expressed by merB (Song et al., Nat Biotechnol. 21:914-919, 2003; International patent PCT/KR02/01934). Likewise, another method was reported that a transgenic plant could be developed by expressing a MRP-type ABC carrier of yeast in a plant when it was over-expressed by merB (Korean Patent No. 0480843).
However, these transgenic plants including a gene with heavy metal resistances outgrew a wild-type one in a contaminated soil but did not have much improved accumulation in the shoot region. In general, a plant for environment purification should not only have resistances against contamination materials but also be able to carry them to the shoot region and thereby, accumulate them there. The reason is that the shoot region of a plant can be more safely and economically harvested and disposed rather than the root.
However, a conventional transgenic plant including a gene with heavy metal resistance a little outgrew a wild-type one in a contaminated soil but turned out to be not be able to effectively pump up heavy metals from root to shoot region and thereby, accumulate them in the shoot region.
In general, a plant uptakes various contamination materials in the environment when it uptakes water. Accordingly, when a plant can uptake more water and transpire it, it can accumulate more contamination materials in its body faster. Therefore, this kind of a plant can decrease contamination in the environment in a shorter term than a wild-type one, contributing to diminishing time and cost for purifying the environment.
In addition, water insufficiency is world-widely raised as a serious problem. The world already started to be desertificated in several areas. This is subsequently causing another serious problem in farming and environment. Therefore, a plant that can need less water and successfully survive in a dry and highly-salted environment should be desperately developed. In particular, when a plant is developed for economically purifying environment in a dry area, it should ideally have improved resistance against drought as well as contamination materials. On the other hand, when a plant can decrease transpiration, it can contribute to purifying environment and increasing agricultural productivity in a very arid area.
Unlike a plant for purifying environment, a crop is required to uptake contamination materials to a minimum. If a crop absorbs heavy metals or other contamination materials, it can hurt people and domestic animals consuming it. Therefore, a crop cannot be poisoned but safe from heavy metals by expressing a gene removing a heavy metal therein. In addition, when a gene accumulating a heavy metal in a cell is artificially less expressed or suppressed to oppositely work, a transgenic plant including the gene can have less accumulated heavy metals than a wild-type one, contributing to development of a safer plant.
On the other hand, a gene related to have heavy metal resistance, salt resistance, drought resistance, and the like can be used to rehabilitate an environment. The rehabilitation of environment can be accomplished by recovering nature artificially or naturally destroyed once and rebuilding ecosystem as it used to be. Accordingly, the environment can not only be refreshed but natural resources also can be preserved, so that human can cohabit with other living species. A method of rehabilitating environment includes removal of contamination materials, planting of a resistant plant, reintroduction of extinct animals, and the like. For example, Daegoo arboretum was created by planting a wild plant and a medicinal plant in a landfill used for dumping trash and thereby, changing it into an ecosystem education center and a rest place. In addition, Kwangjoo Ooncheon reservoir used to be contaminated due to domestic sewage but was now rehabilitated into a natural ecosystem park. However, a landfill may release cadmium with leachate. An abandoned mine can be severely arid. Therefore, genes with high resistance against heavy metals and drought can be usefully used for rehabilitating a landfill or an abandoned mine. In addition, a gene with high salt resistance can be used for rehabilitating a reclaimed land with a high salt concentration.